Search

Although hyponatremia and salt wasting are common in patients with HIV/AIDS, the understanding of their contributing factors is limited. HIV viral protein R (Vpr) contributes to HIV-associated nephropathy. To investigate the effects of Vpr on the distal tubules and on the expression level of the Slc12a3 gene, encoding the sodium-chloride cotransporter (which is responsible for sodium reabsorption in distal nephron segments), single-nucleus RNA sequencing was performed on kidney cortices from three wild-type (WT) and three Vpr transgenic (Vpr Tg) mice. The percentage of distal convoluted tubule (DCT) cells was significantly lower in Vpr Tg mice compared with WT mice (P < 0.05); in Vpr Tg mice, Slc12a3 expression was not significantly different in DCT cells. The Pvalb +  DCT1 subcluster had fewer cells in Vpr Tg mice compared with those in WT mice (P < 0.01). Immunohistochemistry revealed fewer Slc12a3 + Pvalb +  DCT1 segments in Vpr Tg mice. Differential gene expression analysis between Vpr Tg and WT samples in the DCT cluster showed down-regulation of the Ier3 gene, which is an inhibitor of apoptosis. The in vitro knockdown of Ier3 by siRNA transfection induced apoptosis in mouse DCT cells. These observations suggest that the salt-wasting effect of Vpr in Vpr Tg mice is likely mediated by Ier3 down-regulation in DCT1 cells and loss of Slc12a3 + Pvalb +  DCT1 segments., Competing Interests: Disclosure Statement None declared., (Copyright © 2024 American Society for Investigative Pathology. All rights reserved.)

HIV disease remains prevalent in the United States and is particularly prevalent in sub-Saharan Africa. Recent investigations revealed that mitochondrial dysfunction in kidney contributes to HIV-associated nephropathy (HIVAN) in Tg26 transgenic mice. We hypothesized that nicotinamide adenine dinucleotide (NAD) deficiency contributes to energetic dysfunction and progressive tubular injury. We investigated metabolomic mechanisms of HIVAN tubulopathy. Tg26 and wild-type (WT) mice were treated with the farnesoid X receptor (FXR) agonist INT-747 or nicotinamide riboside (NR) from 6 to 12 wk of age. Multiomic approaches were used to characterize kidney tissue transcriptomes and metabolomes. Treatment with INT-747 or NR ameliorated kidney tubular injury, as shown by serum creatinine, the tubular injury marker urinary neutrophil-associated lipocalin, and tubular morphometry. Integrated analysis of metabolomic and transcriptomic measurements showed that NAD levels and production were globally downregulated in Tg26 mouse kidneys, especially nicotinamide phosphoribosyltransferase (NAMPT), the rate-limiting enzyme in the NAD salvage pathway. Furthermore, NAD-dependent deacetylase sirtuin3 activity and mitochondrial oxidative phosphorylation activity were lower in ex vivo proximal tubules from Tg26 mouse kidneys compared with those of WT mice. Restoration of NAD levels in the kidney improved these abnormalities. These data suggest that NAD deficiency might be a treatable target for HIVAN. NEW & NOTEWORTHY The study describes a novel investigation that identified nicotinamide adenine dinucleotide (NAD) deficiency in a widely used HIV-associated nephropathy (HIVAN) transgenic mouse model. We show that INT-747, a farnesoid X receptor agonist, and nicotinamide riboside (NR), a precursor of nicotinamide, each ameliorated HIVAN tubulopathy. Multiomic analysis of mouse kidneys revealed that NAD deficiency was an upstream metabolomic mechanism contributing to HIVAN tubulopathy.

HIV disease remains prevalent in the USA and chronic kidney disease remains a major cause of morbidity in HIV-1-positive patients. Host double-stranded RNA (dsRNA)-activated protein kinase (PKR) is a sensor for viral dsRNA, including HIV-1. We show that PKR inhibition by compound C16 ameliorates the HIV-associated nephropathy (HIVAN) kidney phenotype in the Tg26 transgenic mouse model, with reversal of mitochondrial dysfunction. Combined analysis of single-nucleus RNA-seq and bulk RNA-seq data revealed that oxidative phosphorylation was one of the most downregulated pathways and identified signal transducer and activator of transcription (STAT3) as a potential mediating factor. We identified in Tg26 mice a novel proximal tubular cell cluster enriched in mitochondrial transcripts. Podocytes showed high levels of HIV-1 gene expression and dysregulation of cytoskeleton-related genes, and these cells dedifferentiated. In injured proximal tubules, cell-cell interaction analysis indicated activation of the pro-fibrogenic PKR-STAT3-platelet-derived growth factor (PDGF)-D pathway. These findings suggest that PKR inhibition and mitochondrial rescue are potential novel therapeutic approaches for HIVAN., Competing Interests: TY, KL, AR, BS, KM, YI, YZ, SS, BJ, XY, XW, VT, PS, ML, KO, CW, JK No competing interests declared

Spatial -OMICS technologies facilitate the interrogation of molecular profiles in the context of the underlying histopathology and tissue microenvironment. Paired analysis of histopathology and molecular data can provide pathologists with otherwise unobtainable insights into biological mechanisms. To connect the disparate molecular and histopathologic features into a single workspace, we developed FUSION ( F unctional U nit S tate I dentificati ON in WSIs [Whole Slide Images]), a web-based tool that provides users with a broad array of visualization and analytical tools including deep learning-based algorithms for in-depth interrogation of spatial -OMICS datasets and their associated high-resolution histology images. FUSION enables end-to-end analysis of functional tissue units (FTUs), automatically aggregating underlying molecular data to provide a histopathology-based medium for analyzing healthy and altered cell states and driving new discoveries using "pathomic" features. We demonstrate FUSION using 10x Visium spatial transcriptomics (ST) data from both formalin-fixed paraffin embedded (FFPE) and frozen prepared datasets consisting of healthy and diseased tissue. Through several use-cases, we demonstrate how users can identify spatial linkages between quantitative pathomics, qualitative image characteristics, and spatial --omics.

Background: Mechanisms of progression of diabetic kidney disease (DKD) are not completely understood. This study uses untargeted and targeted mass spectrometry-based proteomics in two independent cohorts on two continents to decipher the mechanisms of DKD in patients with type 2 diabetes., Methods: We conducted untargeted mass spectrometry on urine samples collected at the time of kidney biopsy from Korean patients with type 2 diabetes and biopsy-proven diabetic nephropathy at Seoul National University Hospital (SNUH-DN cohort; n = 64). These findings were validated using targeted mass spectrometry in urine samples from a Chronic Renal Insufficiency Cohort subgroup with type 2 diabetes and DKD (CRIC-T2D; n = 282). Urinary biomarkers/pathways associated with kidney disease progression (doubling of serum creatinine, ≥50% decrease in estimated glomerular filtration rates, or the development of end-stage kidney disease) were identified., Results: SNUH-DN patients had an estimated glomerular filtration rate (eGFR) of 55 mL/min/1.73 m 2 (interquartile range [IQR], 44-75) and random urine protein-to-creatinine ratio of 3.1 g/g (IQR, 1.7-7.0). Urine proteins clustered into two groups, with cluster 2 having a 4.6-fold greater hazard (95% confidence interval [CI], 1.9-11.5) of disease progression than cluster 1 in multivariable-adjusted, time-to-event analyses. Proteins in cluster 2 mapped to 10 pathways, four of the top five of which were complement or complement-related. A high complement score, constructed from urine complement protein abundance, was strongly correlated to 4 of 5 histopathologic DN features and was associated with a 2.4-fold greater hazard (95% CI, 1.0-5.4) of disease progression than a low complement score. Targeted mass spectrometry of the CRIC-T2D participants, who had an eGFR of 42 mL/min/1.73 m 2 (IQR, 37-49) and 24-hr urine protein of 0.48 g (IQR, 0.10-1.87), showed that the complement score similarly segregated them into rapid and slow DKD progression groups. In both cohorts, the complement score had a linear association with disease progression., Conclusions: Urinary proteomic profiling confirms the association between the complement pathway and rapid DKD progression in two independent cohorts. These results suggest a need to further investigate complement pathway inhibition as a novel treatment for DKD.

Many data resources generate, process, store, or provide kidney related molecular, pathological, and clinical data. Reference ontologies offer an opportunity to support knowledge and data integration. The Kidney Precision Medicine Project (KPMP) team contributed to the representation and addition of 329 kidney phenotype terms to the Human Phenotype Ontology (HPO), and identified many subcategories of acute kidney injury (AKI) or chronic kidney disease (CKD). The Kidney Tissue Atlas Ontology (KTAO) imports and integrates kidney-related terms from existing ontologies (e.g., HPO, CL, and Uberon) and represents 259 kidney-related biomarkers. We have also developed a precision medicine metadata ontology (PMMO) to integrate 50 variables from KPMP and CZ CellxGene data resources and applied PMMO for integrative kidney data analysis. The gene expression profiles of kidney gene biomarkers were specifically analyzed under healthy control or AKI/CKD disease states. This work demonstrates how ontology-based approaches support multi-domain data and knowledge integration in precision medicine.

HistoLens is an open-source graphical user interface developed using MATLAB AppDesigner for visual and quantitative analysis of histological datasets. HistoLens enables users to interrogate sets of digitally annotated whole slide images to efficiently characterize histological differences between disease and experimental groups. Users can dynamically visualize the distribution of 448 hand-engineered features quantifying color, texture, morphology, and distribution across microanatomic sub-compartments. Additionally, users can map differentially detected image features within the images by highlighting affected regions. We demonstrate the utility of HistoLens to identify hand-engineered features that correlate with pathognomonic renal glomerular characteristics distinguishing diabetic nephropathy and amyloid nephropathy from the histologically unremarkable glomeruli in minimal change disease. Additionally, we examine the use of HistoLens for glomerular feature discovery in the Tg26 mouse model of HIV-associated nephropathy. We identify numerous quantitative glomerular features distinguishing Tg26 transgenic mice from wild-type mice, corresponding to a progressive renal disease phenotype. Thus, we demonstrate an off-the-shelf and ready-to-use toolkit for quantitative renal pathology applications., (© 2024. The Author(s).)

Chronic kidney disease progresses through the replacement of functional tissue compartments with fibrosis, a maladaptive repair process. Shifting kidney repair toward a physiologically intact architecture, rather than fibrosis, is key to blocking chronic kidney disease progression. Much research into the mechanisms of fibrosis is performed in rodent models with less attention to the human genetic context. Recently, human induced pluripotent stem cell (iPSC)-derived organoids have shown promise in overcoming the limitation. In this study, we developed a fibrosis model that uses human iPSC-based 3-dimensional renal organoids, in which exogenous transforming growth factor-β1 (TGF-β1) induced the production of extracellular matrix. TGF-β1-treated organoids showed tubulocentric collagen 1α1 production by regulating downstream transcriptional regulators, Farnesoid X receptor, phosphorylated mothers against decapentaplegic homolog 3 (p-SMAD3), and transcriptional coactivator with PDZ-binding motif (TAZ). Increased nuclear TAZ expression was confirmed in the tubular epithelium in human kidney biopsies with tubular injury and early fibrosis. A dual bile acid receptor agonist (INT-767) increased Farnesoid X receptor and reduced p-SMAD3 and TAZ, attenuating TGF-β1-induced fibrosis in kidney organoids. Finally, we show that TAZ interacted with TEA-domain transcription factors and p-SMAD3 with TAZ and TEA-domain transcription factor 4 coregulating collagen 1α1 gene transcription. In summary, we establish a novel, readily manipulable fibrogenesis model and posit a role for bile acid receptor agonism early in renal parenchymal fibrosis., (Copyright © 2024. Published by Elsevier Inc.)

Sodium-glucose cotransporter, type 2 inhibitors (SGLT2i) are emerging as the gold standard for treatment of type 2 diabetes (T2D) with renal protective benefits independent of glucose lowering. We took a high-level approach to evaluate the effects of the SGLT2i, empagliflozin (EMPA) on renal metabolism and function in a prediabetic model of metabolic syndrome. Male and female 12-wk-old TallyHo (TH) mice, and their closest genetic lean strain (Swiss-Webster, SW) were treated with a high-milk-fat diet (HMFD) plus/minus EMPA (@0.01%) for 12-wk. Kidney weights and glomerular filtration rate were slightly increased by EMPA in the TH mice. Glomerular feature analysis by unsupervised clustering revealed sexually dimorphic clustering, and one unique cluster relating to EMPA. Periodic acid Schiff (PAS) positive areas, reflecting basement membranes and mesangium were slightly reduced by EMPA. Phasor-fluorescent life-time imaging (FLIM) of free-to-protein bound NADH in cortex showed a marginally greater reliance on oxidative phosphorylation with EMPA. Overall, net urine sodium, glucose, and albumin were slightly increased by EMPA. In TH, EMPA reduced the sodium phosphate cotransporter, type 2 (NaPi-2), but increased sodium hydrogen exchanger, type 3 (NHE3). These changes were absent or blunted in SW. EMPA led to changes in urine exosomal microRNA profile including, in females, enhanced levels of miRs 27a-3p, 190a-5p, and 196b-5p. Network analysis revealed "cancer pathways" and "FOXO signaling" as the major regulated pathways. Overall, EMPA treatment to prediabetic mice with limited renal disease resulted in modifications in renal metabolism, structure, and transport, which may preclude and underlie protection against kidney disease with developing T2D. NEW & NOTEWORTHY Renal protection afforded by sodium glucose transporter, type 2 inhibitors (SGLT2i), e.g., empagliflozin (EMPA) involves complex intertwined mechanisms. Using a novel mouse model of obesity with insulin resistance, the TallyHo/Jng (TH) mouse on a high-milk-fat diet (HMFD), we found subtle changes in metabolism including altered regulation of sodium transporters that line the renal tubule. New potential epigenetic determinants of metabolic changes relating to FOXO and cancer signaling pathways were elucidated from an altered urine exosomal microRNA signature.

Artificial intelligence (AI) has extensive applications in a wide range of disciplines including healthcare and clinical practice. Advances in high-resolution whole-slide brightfield microscopy allow for the digitization of histologically stained tissue sections, producing gigapixel-scale whole-slide images (WSI). The significant improvement in computing and revolution of deep neural network (DNN)-based AI technologies over the last decade allow us to integrate massively parallelized computational power, cutting-edge AI algorithms, and big data storage, management, and processing. Applied to WSIs, AI has created opportunities for improved disease diagnostics and prognostics with the ultimate goal of enhancing precision medicine and resulting patient care. The National Institutes of Health (NIH) has recognized the importance of developing standardized principles for data management and discovery for the advancement of science and proposed the Findable, Accessible, Interoperable, Reusable, (FAIR) Data Principles 1 with the goal of building a modernized biomedical data resource ecosystem to establish collaborative research communities. In line with this mission and to democratize AI-based image analysis in digital pathology, we propose ComPRePS: an end-to-end automated Comp utational Re nal P athology S uite which combines massive scalability, on-demand cloud computing, and an easy-to-use web-based user interface for data upload, storage, management, slide-level visualization, and domain expert interaction. Moreover, our platform is equipped with both in-house and collaborator developed sophisticated AI algorithms in the back-end server for image analysis to identify clinically relevant micro-anatomic functional tissue units (FTU) and to extract image features.

A gradual decline in renal function occurs even in healthy aging individuals. In addition to aging, per se, concurrent metabolic syndrome and hypertension, which are common in the aging population, can induce mitochondrial dysfunction and inflammation, which collectively contribute to age-related kidney dysfunction and disease. This study examined the role of the nuclear hormone receptors, the estrogen-related receptors (ERRs), in regulation of age-related mitochondrial dysfunction and inflammation. The ERRs were decreased in both aging human and mouse kidneys and were preserved in aging mice with lifelong caloric restriction (CR). A pan-ERR agonist, SLU-PP-332, was used to treat 21-month-old mice for 8 weeks. In addition, 21-month-old mice were treated with a stimulator of interferon genes (STING) inhibitor, C-176, for 3 weeks. Remarkably, similar to CR, an 8-week treatment with a pan-ERR agonist reversed the age-related increases in albuminuria, podocyte loss, mitochondrial dysfunction, and inflammatory cytokines, via the cyclic GMP-AMP synthase-STING and STAT3 signaling pathways. A 3-week treatment of 21-month-old mice with a STING inhibitor reversed the increases in inflammatory cytokines and the senescence marker, p21/cyclin dependent kinase inhibitor 1A (Cdkn1a), but also unexpectedly reversed the age-related decreases in PPARG coactivator (PGC)-1α, ERRα, mitochondrial complexes, and medium chain acyl coenzyme A dehydrogenase (MCAD) expression. These studies identified ERRs as CR mimetics and as important modulators of age-related mitochondrial dysfunction and inflammation. These findings highlight novel druggable pathways that can be further evaluated to prevent progression of age-related kidney disease., Competing Interests: Disclosure Statement None declared., (Copyright © 2023 American Society for Investigative Pathology. Published by Elsevier Inc. All rights reserved.)

Farnesoid X receptor (FXR) activation reduces renal inflammation, but the underlying mechanisms remain elusive. Neutrophil extracellular traps (NETs) are webs of DNA formed when neutrophils undergo specialized programmed cell death (NETosis). The signaling lipid sphingosine-1-phosphate (S1P) stimulates NETosis via its receptor on neutrophils. Here, we identify FXR as a negative regulator of NETosis via repressing S1P signaling. We determined the effects of the FXR agonist obeticholic acid (OCA) in mouse models of adenosine phosphoribosyltransferase (APRT) deficiency and Alport syndrome, both genetic disorders that cause chronic kidney disease. Renal FXR activity is greatly reduced in both models, and FXR agonism reduces disease severity. Renal NETosis and sphingosine kinase 1 ( Sphk1 ) expression are increased in diseased mice, and they are reduced by OCA in both models. Genetic deletion of FXR increases Sphk1 expression, and Sphk1 expression correlates with NETosis. Importantly, kidney S1P levels in Alport mice are two-fold higher than controls, and FXR agonism restores them back to baseline. Short-term inhibition of sphingosine synthesis in Alport mice with severe kidney disease reverses NETosis, establishing a causal relationship between S1P signaling and renal NETosis. Finally, extensive NETosis is present in human Alport kidney biopsies (six male, nine female), and NETosis severity correlates with clinical markers of kidney disease. This suggests the potential clinical relevance of the newly identified FXR-S1P-NETosis pathway. In summary, FXR agonism represses kidney Sphk1 expression. This inhibits renal S1P signaling, thereby reducing neutrophilic inflammation and NETosis. NEW & NOTEWORTHY Many preclinical studies have shown that the farnesoid X receptor (FXR) reduces renal inflammation, but the mechanism is poorly understood. This report identifies FXR as a novel regulator of neutrophilic inflammation and NETosis via the inhibition of sphingosine-1-phosphate signaling. Additionally, NETosis severity in human Alport kidney biopsies correlates with clinical markers of kidney disease. A better understanding of this signaling axis may lead to novel treatments that prevent renal inflammation and chronic kidney disease.

Background: Pregnancy complications are risk factors for cardiovascular disease (CVD). Little is known about the role of renal biomarkers measured shortly after delivery, individually or in combination with pregnancy complications, in predicting subsequent severe maternal CVD., Methods and Results: This study included 566 mothers of diverse races and ethnicities from the Boston Birth cohort, enrolled at delivery and followed prospectively. Plasma creatinine and CysC (cystatin C) were measured 1 to 3 days after delivery. CVD during follow-up was defined by physician diagnoses in electronic medical records. Associations of renal biomarkers and pregnancy complications with time-to-CVD events were assessed using Cox proportional hazards models. During an average of 10.3±3.2 years of follow-up, 30 mothers developed 1 or more CVDs. Only a modest association was observed between creatinine and risk of CVD. In comparison, we found that per 0.1 mg/L increase of CysC was associated with a hazard ratio (HR) of 1.2 (95% CI, 1.1-1.4) for CVD after adjusting for covariates. Compared with those without preeclampsia and with normal CysC level (≤75th percentile), mothers with preeclampsia and elevated CysC (>75th percentile) had the highest risk of CVD (HR, 4.6 [95% CI, 1.7-17.7]), whereas mothers with preeclampsia only or with elevated CysC only did not have significantly increased CVD risk. Similar synergistic effects for CVD were observed between CysC and preterm delivery., Conclusions: In this sample of US, traditionally underrepresented multiracial and multiethnic high-risk mothers, elevated maternal plasma CysC, independently and jointly with pregnancy complications, increased risk of CVD later in life. These findings warrant further investigation., Registration: URL: https://www.clinicaltrials.gov; Unique identifier: NCT03228875.

Autosomal recessive polycystic kidney disease (ARPKD; MIM#263200) is a severe, hereditary, hepato-renal fibrocystic disorder that leads to early childhood morbidity and mortality. Typical forms of ARPKD are caused by pathogenic variants in the PKHD1 gene, which encodes the fibrocystin/polyductin (FPC) protein. MYC overexpression has been proposed as a driver of renal cystogenesis, but little is known about MYC expression in recessive PKD. In the current study, we provide the first evidence that MYC is overexpressed in kidneys from ARPKD patients and confirm that MYC is upregulated in cystic kidneys from cpk mutant mice. In contrast, renal MYC expression levels were not altered in several Pkhd1 mutant mice that lack a significant cystic kidney phenotype. We leveraged previous observations that the carboxy-terminus of mouse FPC (FPC-CTD) is proteolytically cleaved through Notch-like processing, translocates to the nucleus, and binds to double stranded DNA, to examine whether the FPC-CTD plays a role in regulating MYC/Myc transcription. Using immunofluorescence, reporter gene assays, and ChIP, we demonstrate that both human and mouse FPC-CTD can localize to the nucleus, bind to the MYC/Myc P1 promoter, and activate MYC/Myc expression. Interestingly, we observed species-specific differences in FPC-CTD intracellular trafficking. Furthermore, our informatic analyses revealed limited sequence identity of FPC-CTD across vertebrate phyla and database queries identified temporal differences in PKHD1 / Pkhd1 and CYS1 / Cys1 expression patterns in mouse and human kidneys. Given that cystin, the Cys1 gene product, is a negative regulator of Myc transcription, these temporal differences in gene expression could contribute to the relative renoprotection from cystogenesis in Pkhd1 -deficient mice. Taken together, our findings provide new mechanistic insights into differential mFPC-CTD and hFPC-CTD regulation of MYC expression in renal epithelial cells, which may illuminate the basis for the phenotypic disparities between human patients with PKHD1 pathogenic variants and Pkhd1 -mutant mice., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2023 Harafuji, Yang, Wu, Thiruvengadam, Gordish-Dressman, Thompson, Bell, Rosenberg, Dafinger, Liebau, Bebok, Caldovic and Guay-Woodford.)

Chronic pancreatitis results in the formation of pancreatic intraepithelial neoplasia (PanIN) and poses a risk of developing pancreatic cancer. Our previous study demonstrated that Krüppel-like factor 5 (KLF5) is necessary for forming acinar-to-ductal metaplasia (ADM) in acute pancreatitis. Here, we investigated the role of KLF5 in response to chronic injury in the pancreas. Human tissues originating from chronic pancreatitis patients showed increased levels of epithelial KLF5. An inducible genetic model combining the deletion of Klf5 and the activation of Kras G12D mutant expression in pancreatic acinar cells together with chemically induced chronic pancreatitis was used. The chronic injury resulted in increased levels of KLF5 in both control and Kras G12D mutant mice. Furthermore, it led to numerous ADM and PanIN lesions and extensive fibrosis in the KRAS mutant mice. In contrast, pancreata with Klf5 loss (with or without Kras G12D ) failed to develop ADM, PanIN, or significant fibrosis. Furthermore, the deletion of Klf5 reduced the expression level of cytokines and fibrotic components such as Il1b , Il6 , Tnf , Tgfb1 , Timp1 , and Mmp9 . Notably, using ChIP-PCR, we showed that KLF5 binds directly to the promoters of Il1b , Il6 , and Tgfb1 genes. In summary, the inactivation of Klf5 inhibits ADM and PanIN formation and the development of pancreatic fibrosis.

Rationale & Objective: Patients hospitalized with COVID-19 are at increased risk for major adverse kidney events (MAKE). We sought to identify plasma biomarkers predictive of MAKE in patients hospitalized with COVID-19., Study Design: Prospective cohort study., Setting & Participants: A total of 576 patients hospitalized with COVID-19 between March 2020 and January 2021 across 3 academic medical centers., Exposure: Twenty-six plasma biomarkers of injury, inflammation, and repair from first available blood samples collected during hospitalization., Outcome: MAKE, defined as KDIGO stage 3 acute kidney injury (AKI), dialysis-requiring AKI, or mortality up to 60 days., Analytical Approach: Cox proportional hazards regression to associate biomarker level with MAKE. We additionally applied the least absolute shrinkage and selection operator (LASSO) and random forest regression for prediction modeling and estimated model discrimination with time-varying C index., Results: The median length of stay for COVID-19 hospitalization was 9 (IQR, 5-16) days. In total, 95 patients (16%) experienced MAKE. Each 1 SD increase in soluble tumor necrosis factor receptor 1 (sTNFR1) and sTNFR2 was significantly associated with an increased risk of MAKE (adjusted HR [AHR], 2.30 [95% CI, 1.86-2.85], and AHR, 2.26 [95% CI, 1.73-2.95], respectively). The C index of sTNFR1 alone was 0.80 (95% CI, 0.78-0.84), and the C index of sTNFR2 was 0.81 (95% CI, 0.77-0.84). LASSO and random forest regression modeling using all biomarkers yielded C indexes of 0.86 (95% CI, 0.83-0.89) and 0.84 (95% CI, 0.78-0.91), respectively., Limitations: No control group of hospitalized patients without COVID-19., Conclusions: We found that sTNFR1 and sTNFR2 are independently associated with MAKE in patients hospitalized with COVID-19 and can both also serve as predictors for adverse kidney outcomes., Plain-Language Summary: Patients hospitalized with COVID-19 are at increased risk for long-term adverse health outcomes, but not all patients suffer long-term kidney dysfunction. Identification of patients with COVID-19 who are at high risk for adverse kidney events may have important implications in terms of nephrology follow-up and patient counseling. In this study, we found that the plasma biomarkers soluble tumor necrosis factor receptor 1 (sTNFR1) and sTNFR2 measured in hospitalized patients with COVID-19 were associated with a greater risk of adverse kidney outcomes. Along with clinical variables previously shown to predict adverse kidney events in patients with COVID-19, both sTNFR1 and sTNFR2 are also strong predictors of adverse kidney outcomes., (Copyright © 2023 National Kidney Foundation, Inc. Published by Elsevier Inc. All rights reserved.)

Age is a significant risk factor for the coronavirus disease 2019 (COVID-19) severity due to immunosenescence and certain age-dependent medical conditions (e.g., obesity, cardiovascular disorder, and chronic respiratory disease). However, despite the well-known influence of age on autoantibody biology in health and disease, its impact on the risk of developing severe COVID-19 remains poorly explored. Here, we performed a cross-sectional study of autoantibodies directed against 58 targets associated with autoimmune diseases in 159 individuals with different COVID-19 severity (71 mild, 61 moderate, and 27 with severe symptoms) and 73 healthy controls. We found that the natural production of autoantibodies increases with age and is exacerbated by SARS-CoV-2 infection, mostly in severe COVID-19 patients. Multiple linear regression analysis showed that severe COVID-19 patients have a significant age-associated increase of autoantibody levels against 16 targets (e.g., amyloid β peptide, β catenin, cardiolipin, claudin, enteric nerve, fibulin, insulin receptor a, and platelet glycoprotein). Principal component analysis with spectrum decomposition and hierarchical clustering analysis based on these autoantibodies indicated an age-dependent stratification of severe COVID-19 patients. Random forest analysis ranked autoantibodies targeting cardiolipin, claudin, and platelet glycoprotein as the three most crucial autoantibodies for the stratification of severe COVID-19 patients ≥50 years of age. Follow-up analysis using binomial logistic regression found that anti-cardiolipin and anti-platelet glycoprotein autoantibodies significantly increased the likelihood of developing a severe COVID-19 phenotype with aging. These findings provide key insights to explain why aging increases the chance of developing more severe COVID-19 phenotypes., (© 2023. Springer Nature Limited.)

Diabetes mellitus is the leading cause of cardiovascular and renal disease in the United -States. Despite the beneficial interventions available for patients with diabetes, there remains a need for additional therapeutic targets and therapies in diabetic kidney disease (DKD). Inflammation and oxidative stress are increasingly recognized as important causes of renal diseases. Inflammation is closely associated with mitochondrial damage. The molecular connection between inflammation and mitochondrial metabolism remains to be elucidated. Recently, nicotinamide adenine nucleotide (NAD+) metabolism has been found to regulate immune function and inflammation. In the present studies, we tested the hypothesis that enhancing NAD metabolism could prevent inflammation in and progression of DKD. We found that treatment of db/db mice with type 2 diabetes with nicotinamide riboside (NR) prevented several manifestations of kidney dysfunction (i.e., albuminuria, increased urinary kidney injury marker-1 (KIM1) excretion, and pathologic changes). These effects were associated with decreased inflammation, at least in part via inhibiting the activation of the cyclic GMP-AMP synthase-stimulator of interferon genes (cGAS-STING) signaling pathway. An antagonist of the serum stimulator of interferon genes (STING) and whole-body STING deletion in diabetic mice showed similar renoprotection. Further analysis found that NR increased SIRT3 activity and improved mitochondrial function, which led to decreased mitochondrial DNA damage, a trigger for mitochondrial DNA leakage which activates the cGAS-STING pathway. Overall, these data show that NR supplementation boosted NAD metabolism to augment mitochondrial function, reducing inflammation and thereby preventing the progression of diabetic kidney disease., Competing Interests: Conflict of interest The authors declare that they have no conflicts of interest with the contents of this article., (Copyright © 2023 The Authors. Published by Elsevier Inc. All rights reserved.)

Cell-specific microRNA (miRNA) expression estimates are important in characterizing the localization of miRNA signaling within tissues. Much of these data are obtained from cultured cells, a process known to significantly alter miRNA expression levels. Thus, our knowledge of in vivo cell miRNA expression estimates is poor. We previously demonstrated expression microdissection-miRNA-sequencing (xMD-miRNA-seq) to acquire in vivo estimates, directly from formalin-fixed tissues, albeit with a limited yield. In this study, we optimized each step of the xMD process, including tissue retrieval, tissue transfer, film preparation, and RNA isolation, to increase RNA yields and ultimately show strong enrichment for in vivo miRNA expression by qPCR array. These method improvements, such as the development of a noncrosslinked ethylene vinyl acetate membrane, resulted in a 23- to 45-fold increase in miRNA yield, depending on the cell type. By qPCR, miR-200a increased by 14-fold in xMD-derived small intestine epithelial cells, with a concurrent 336-fold reduction in miR-143 relative to the matched nondissected duodenal tissue. xMD is now an optimized method to obtain robust in vivo miRNA expression estimates from cells. xMD will allow formalin-fixed tissues from surgical pathology archives to make theragnostic biomarker discoveries., (Copyright © 2023 United States & Canadian Academy of Pathology. Published by Elsevier Inc. All rights reserved.)

Objective: It is suggested that hidradenitis suppurativa (HS) is more prevalent and causes greater morbidity in Black patients than in White. Clinical data are however lacking., Patients and Methods: We therefore describe HS risk factors, disease severity and clinical phenotypes in the Blacks and Whites. Patients referred for HS between 1984 and 2019 at the Johns Hopkins Hospital were identified using the Pathology Data System (PDS). Clinical and sociodemographic characteristics were extracted and the van der Zee & Jemec HS clinical phenotypes were recovered., Results: A total of 278 patients were identified. Ethnically, 108 (38.8%) were White, and 170 (61.2%) Black. The following HS phenotypes were found: Regular (n=193, 69.4%), scarring folliculitis (n=40, 1.4%) frictional furuncle (11.2%), conglobata (n=9, 3.2%), syndromic (n=3, 1.1%) and ectopic (n=2, 0.7%). No statistically significant ethnic differences in clinical presentation were found. Blacks however had more severe diseases than Whites (p= 0.024 for trend). With multivariate logistic regression analysis, we found that male sex, disease duration, and smoking were independent predictors of regular HS phenotype. Major limitations are the limited number of cases studied and the lack of data regarding response to therapies., Conclusions: Demographics and phenotypical presentation of HS patients do not seem to be associated with Black ethnicity. However, there is a significant trend for Blacks to present with more Hurley stage 2 and 3 disease compared to White patients. It is speculated that ethnic differences are epiphenomena to social factors, highlighting the broader importance of ethnicity.

The major goals of the Kidney Precision Medicine Project (KPMPP) are to establish a molecular atlas of the kidney in health and disease and improve our understanding of the molecular drivers of CKD and AKI. In this clinical-pathologic-molecular correlation, we describe the case of a 38-year-old woman without any history of CKD who underwent a research kidney biopsy in the setting of AKI suspected to be due to nonsteroidal anti-inflammatory use after cesarean section delivery. The participant's histopathology was consistent with mild acute tubular injury, without significant interstitial fibrosis or tubular atrophy. This diagnosis was supported by analysis of the glomerular and tubulointerstitial proteomes. The proteomic interrogation revealed a molecular landscape that demonstrated differences in kidney prostaglandin synthesis that may be in response to nonsteroidal anti-inflammatory drugs and signs of intrarenal inflammation and fibrosis that were not evident by histopathology alone., (Copyright © 2022 by the American Society of Nephrology.)

Background: COVID-19 is a heterogenous disease resulting in long-term sequela in predisposed individuals. It is not uncommon that recovering patients endure non-respiratory ill-defined manifestations, including anosmia, and neurological and cognitive deficit persisting beyond recovery-a constellation of conditions that are grouped under the umbrella of long-term COVID-19 syndrome. Association between COVID-19 and autoimmune responses in predisposed individuals was shown in several studies., Aim and Methods: To investigate autoimmune responses against neuronal and CNS autoantigens in SARS-CoV-2-infected patients, we performed a cross-sectional study with 246 participants, including 169 COVID-19 patients and 77 controls. Levels of antibodies against the acetylcholine receptor, glutamate receptor, amyloid β peptide, alpha-synucleins, dopamine 1 receptor, dopamine 2 receptor, tau protein, GAD-65, N-methyl D-aspartate (NMDA) receptor, BDNF, cerebellar, ganglioside, myelin basic protein, myelin oligodendrocyte glycoprotein, S100-B, glial fibrillary acidic protein, and enteric nerve were measured using an Enzyme-Linked Immunosorbent Assay (ELISA). Circulating levels of autoantibodies were compared between healthy controls and COVID-19 patients and then classified by disease severity (mild [ n = 74], severe [ n = 65], and requiring supplemental oxygen [ n = 32])., Results: COVID-19 patients were found to have dysregulated autoantibody levels correlating with the disease severity, e.g., IgG to dopamine 1 receptor, NMDA receptors, brain-derived neurotrophic factor, and myelin oligodendrocyte glycoprotein. Elevated levels of IgA autoantibodies against amyloid β peptide, acetylcholine receptor, dopamine 2 receptor, myelin basic protein, and α-synuclein were detected in COVID-19 patients compared with healthy controls. Lower IgA autoantibody levels against NMDA receptors, and IgG autoantibodies against glutamic acid decarboxylase 65, amyloid β peptide, tau protein, enteric nerve, and S100-B were detected in COVID-19 patients versus healthy controls. Some of these antibodies have known clinical correlations with symptoms commonly reported in the long COVID-19 syndrome., Conclusions: Overall, our study shows a widespread dysregulation in the titer of various autoantibodies against neuronal and CNS-related autoantigens in convalescent COVID-19 patients. Further research is needed to provide insight into the association between these neuronal autoantibodies and the enigmatic neurological and psychological symptoms reported in COVID-19 patients., Competing Interests: Aristo Vojdani is the co-owner, CEO and technical director of Immunosciences Lab., Inc. in Los Angeles, CA. He is the Chief Scientific Advisor in a consultancy position for Cyrex Labs, LLC in Phoenix, AZ. The rest of authors declare no conflict of interest.

The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection is associated with increased levels of autoantibodies targeting immunological proteins such as cytokines and chemokines. Reports further indicate that COVID-19 patients may develop a broad spectrum of autoimmune diseases due to reasons not fully understood. Even so, the landscape of autoantibodies induced by SARS-CoV-2 infection remains uncharted territory. To gain more insight, we carried out a comprehensive assessment of autoantibodies known to be linked to diverse autoimmune diseases observed in COVID-19 patients in a cohort of 231 individuals, of which 161 were COVID-19 patients (72 with mild, 61 moderate, and 28 with severe disease) and 70 were healthy controls. Dysregulated IgG and IgA autoantibody signatures, characterized mainly by elevated concentrations, occurred predominantly in patients with moderate or severe COVID-19 infection. Autoantibody levels often accompanied anti-SARS-CoV-2 antibody concentrations while stratifying COVID-19 severity as indicated by random forest and principal component analyses. Furthermore, while young versus elderly COVID-19 patients showed only slight differences in autoantibody levels, elderly patients with severe disease presented higher IgG autoantibody concentrations than young individuals with severe COVID-19. This work maps the intersection of COVID-19 and autoimmunity by demonstrating the dysregulation of multiple autoantibodies triggered during SARS-CoV-2 infection. Thus, this cross-sectional study suggests that SARS-CoV-2 infection induces autoantibody signatures associated with COVID-19 severity and several autoantibodies that can be used as biomarkers of COVID-19 severity, indicating autoantibodies as potential therapeutical targets for these patients., (© 2023 The Authors. Journal of Medical Virology published by Wiley Periodicals LLC.)

In patients with severe #COVID19, increased levels of autoantibodies against PAR1 were found. These might serve as allosteric agonists of PAR1 on endothelial cells and platelets, and thus might contribute to the pathogenesis of microthrombosis in COVID-19. https://bit.ly/3pqM9Vv., Competing Interests: Conflict of Interest: T. Bahmer reports grants from BMBF (unrestricted research grant for the German Center for Lung Research (DZL) and National Pandemic Cohort Network (NAPKON) – Coordinating Study Site for population-based cohort platform); lecture fees from Novartis, AstraZeneca, and Chiesi; support for attending the American Thoracic Society Conference from Chiesi; and advisory board participation with GlaxoSmithKline, Boehringer Ingelheim, Roche and AstraZeneca, outside the submitted work. D. Frank reports grants from DFG (467267736), BMBF and DZHK; consulting fees and support for attending meetings from Edwards Lifesciences and Medtronic; lecture honoraria from Edwards Lifesciences, Medtronic, Astra Zeneca, Pfizer, BMS, Novartis, Bayer and Abbott; and participation on advisory boards with BMS, Boehringer Ingelheim, Daiichi Sankyo, outside the submitted work. A.Z. Rosenberg reports grants from the NIH (NIDDK and NHLBI) outside the submitted work. CellTrend is owned by K. Schulze-Forster; CellTrend produces ELISA kits for the determination of antibodies against GPCR. G. Riemekasten reports consulting fees from CellTrend GmbH outside the submitted work. S. Schreiber reports consulting fees from Abbvie, Arena, BMS, Biogen, Celltrion, Celgene, IMAB, Gilead, MSD, Mylan, Pfizer, Fresenius, Janssen, Takeda, Theravance, Prevention Bio, Protagonist and Falk, outside the submitted work. All other authors have nothing to disclose., (Copyright ©The authors 2022.)

The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) spike (S) protein binds angiotensin-converting enzyme 2 as its primary infection mechanism. Interactions between S and endogenous proteins occur after infection but are not well understood. We profiled binding of S against >9000 human proteins and found an interaction between S and human estrogen receptor α (ERα). Using bioinformatics, supercomputing, and experimental assays, we identified a highly conserved and functional nuclear receptor coregulator (NRC) LXD-like motif on the S2 subunit. In cultured cells, S DNA transfection increased ERα cytoplasmic accumulation, and S treatment induced ER-dependent biological effects. Non-invasive imaging in SARS-CoV-2-infected hamsters localized lung pathology with increased ERα lung levels. Postmortem lung experiments from infected hamsters and humans confirmed an increase in cytoplasmic ERα and its colocalization with S in alveolar macrophages. These findings describe the discovery of a S-ERα interaction, imply a role for S as an NRC, and advance knowledge of SARS-CoV-2 biology and coronavirus disease 2019 pathology.

Background: Genetic variants in apolipoprotein L1 (APOL1), a protein that protects humans from infection with African trypanosomes, explain a substantial proportion of the excess risk of chronic kidney disease affecting individuals with sub-Saharan ancestry. The mechanisms by which risk variants damage kidney cells remain incompletely understood. In preclinical models, APOL1 expressed in podocytes can lead to significant kidney injury. In humans, studies in kidney transplant suggest that the effects of APOL1 variants are predominantly driven by donor genotype. Less attention has been paid to a possible role for circulating APOL1 in kidney injury., Methods: Using liquid chromatography-tandem mass spectrometry, the concentrations of APOL1 were measured in plasma and urine from participants in the Seattle Kidney Study. Asymmetric flow field-flow fractionation was used to evaluate the size of APOL1-containing lipoprotein particles in plasma. Transgenic mice that express wild-type or risk variant APOL1 from an albumin promoter were treated to cause kidney injury and evaluated for renal disease and pathology., Results: In human participants, urine concentrations of APOL1 were correlated with plasma concentrations and reduced kidney function. Risk variant APOL1 was enriched in larger particles. In mice, circulating risk variant APOL1-G1 promoted kidney damage and reduced podocyte density without renal expression of APOL1., Conclusions: These results suggest that plasma APOL1 is dynamic and contributes to the progression of kidney disease in humans, which may have implications for treatment of APOL1-associated kidney disease and for kidney transplantation., Competing Interests: I have read the journal’s policy and the authors of this manuscript have the following competing interests: We appreciate receiving BAC/APOL1 mice from Merck, Kenilworth, NJ. We appreciate receiving APOL1 antibodies from Genentech, South San Francisco, CA. Dr. Ian de Boer reports consulting fees from AstraZeneca, Bayer, Boehringer-Ingelheim, Cyclerion Therapeutics, George Clinical, Gilead, Goldfinch Bio, and Ironwood. This does not alter our adherence to PLOS ONE policies on sharing data and materials. Interested researchers are able to obtain BAC/APOL1 mice from Merck (contact is Lisa Gentile), APOL1 antibodies from Genentech (contact is Suzie Scales, PhD), and Alb/APOL1 transgenic mice from NIDDK, NIH (contact is Jeffrey Kopp, MD), under an appropriate material transfer agreement.

Introduction: The mechanisms in podocytes that mediate the pathologic effects of the APOL1 high-risk (HR) variants remain incompletely understood, although various molecular and cellular mechanisms have been proposed. We previously established conditionally immortalized human urine-derived podocyte-like epithelial cell (HUPEC) lines to investigate APOL1 HR variant-induced podocytopathy., Methods: We conducted comprehensive transcriptomic analysis, including mRNA, microRNA (miRNA), and transfer RNA fragments (tRFs), to characterize the transcriptional profiles in undifferentiated and differentiated HUPEC with APOL1 HR (G1/G2, 2 cell lines) and APOL1 low-risk (LR) (G0/G0, 2 cell lines) genotypes. We reanalyzed single-cell RNA-seq data from urinary podocytes from focal segmental glomerulosclerosis (FSGS) subjects to characterize the effect of APOL1 genotypes on podocyte transcriptomes., Results: Differential expression analysis showed that the ribosomal pathway was one of the most enriched pathways, suggesting that altered function of the translation initiation machinery may contribute to APOL1 variant-induced podocyte injury. Expression of genes related to the elongation initiation factor 2 pathway was also enriched in the APOL1 HR urinary podocytes from single-cell RNA-seq, supporting a prior report on the role of this pathway in APOL1 -associated cell injury. Expression of microRNA and tRFs were analyzed, and the profile of small RNAs differed by both differentiation status and APOL1 genotype., Conclusion: We have profiled the transcriptomic landscape of human podocytes, including mRNA, miRNA, and tRF, to characterize the effects of differentiation and of different APOL1 genotypes. The candidate pathways, miRNAs, and tRFs described here expand understanding of APOL1 -associated podocytopathies.

Background: Cardiovascular magnetic resonance (CMR) is emerging as an important tool for cardiac allograft assessment. Native T1 mapping may add value in identifying rejection and in assessing graft dysfunction and myocardial fibrosis burden. We hypothesized that CMR native T1 values and features of textural analysis of T1 maps would identify acute rejection, and in a secondary analysis, correlate with markers of graft dysfunction, and with fibrosis percentage from endomyocardial biopsy (EMB)., Methods: Fifty cases with simultaneous EMB, right heart catheterization, and 1.5 T CMR with breath-held T1 mapping via modified Look-Locker inversion recovery (MOLLI) in 8 short-axis slices and subsequent quantification of mean and peak native T1 values, were performed on 24 pediatric subjects. A single mid-ventricular slice was used for image texture analysis using nine gray-level co-occurrence matrix features. Digital quantification of Masson trichrome stained EMB samples established degree of fibrosis. Markers of graft dysfunction, including serum brain natriuretic peptide levels and hemodynamic measurements from echocardiography, catheterization, and CMR were collated. Subjects were divided into three groups based on degree of rejection: acute rejection requiring new therapy, mild rejection requiring increased ongoing therapy, and no rejection with no change in treatment. Statistical analysis included student's t-test and linear regression., Results: Peak and mean T1 values were significantly associated with acute rejection, with a monotonic trend observed with increased grade of rejection. Texture analysis demonstrated greater spatial heterogeneity in T1 values, as demonstrated by energy, entropy, and variance, in cases requiring treatment. Interestingly, 2 subjects who required increased therapy despite low grade EMB results had abnormal peak T1 values. Peak T1 values also correlated with increased BNP, right-sided filling pressures, and capillary wedge pressures. There was no difference in histopathological fibrosis percentage among the 3 groups; histopathological fibrosis did not correlate with T1 values or markers of graft dysfunction., Conclusion: In pediatric heart transplant patients, native T1 values identify acute rejection requiring treatment and may identify graft dysfunction. CMR shows promise as an important tool for evaluation of cardiac grafts in children, with T1 imaging outperforming biopsy findings in the assessment of rejection., (© 2022. The Author(s).)

Cross-reactive immunity between SARS-CoV-2 and other related coronaviruses has been well-documented, and it may play a role in preventing severe COVID-19. Epidemiological studies early in the pandemic showed a geographical association between high influenza vaccination rates and lower incidence of SARS-CoV-2 infection. We, therefore, analyzed whether exposure to influenza A virus (IAV) antigens could influence the T cell repertoire in response to SARS-CoV-2, indicating a heterologous immune response between these 2 unrelated viruses. Using artificial antigen-presenting cells (aAPCs) combined with real-time reverse-transcription PCR (RT-qPCR), we developed a sensitive assay to quickly screen for antigen-specific T cell responses and detected a significant correlation between responses to SARS-CoV-2 epitopes and IAV dominant epitope (M158-66). Further analysis showed that some COVID-19 convalescent donors exhibited both T cell receptor (TCR) specificity and functional cytokine responses to multiple SARS-CoV-2 epitopes and M158-66. Utilizing an aAPC-based stimulation/expansion assay, we detected cross-reactive T cells with specificity to SARS-CoV-2 and IAV. In addition, TCR sequencing of the cross-reactive and IAV-specific T cells revealed similarities between the TCR repertoires of the two populations. These results indicate that heterologous immunity shaped by our exposure to other unrelated endemic viruses may affect our immune response to novel viruses such as SARS-CoV-2.

HIV-associated nephropathy (HIVAN) impairs functions of both glomeruli and tubules. Attention has been previously focused on the HIVAN glomerulopathy. Tubular injury has drawn increased attention because sodium wasting is common in hospitalized HIV/AIDS patients. We used viral protein R (Vpr)-transgenic mice to investigate the mechanisms whereby Vpr contributes to urinary sodium wasting. In phosphoenolpyruvate carboxykinase promoter-driven Vpr-transgenic mice, in situ hybridization showed that Vpr mRNA was expressed in all nephron segments, including the distal convoluted tubule. Vpr-transgenic mice, compared with wild-type littermates, markedly increased urinary sodium excretion, despite similar plasma renin activity and aldosterone levels. Kidneys from Vpr-transgenic mice also markedly reduced protein abundance of the Na+-Cl- cotransporter (NCC), while mineralocorticoid receptor (MR) protein expression level was unchanged. In African green monkey kidney cells, Vpr abrogated the aldosterone-mediated stimulation of MR transcriptional activity. Gene expression of Slc12a3 (NCC) in Vpr-transgenic mice was significantly lower compared with wild-type mice, assessed by both qRT-PCR and RNAScope in situ hybridization analysis. Chromatin immunoprecipitation assays identified multiple MR response elements (MRE), located from 5 kb upstream of the transcription start site and extending to the third exon of the SLC12A3 gene. Mutation of MRE and SP1 sites in the SLC12A3 promoter region abrogated the transcriptional responses to aldosterone and Vpr, indicating that functional MRE and SP1 are required for the SLC12A3 gene suppression in response to Vpr. Thus, Vpr attenuates MR transcriptional activity and inhibits Slc12a3 transcription in the distal convoluted tubule and contributes to salt wasting in Vpr-transgenic mice., Competing Interests: Author JBK holds a patent relating to monoclonal antibodies to HIV-1 Vpr and methods of using same. United States Patent 7,993,647 (2015). No other conflicts of interest, financial or otherwise, are declared by the authors. This does not alter our adherence to PLOS ONE policies on sharing data and materials.

Introduction: Proliferative lupus nephritis (LN) progresses to end-stage kidney disease (ESKD) in roughly 10% of the cases despite treatment. Other than achieving <0.8 g/24h proteinuria at 12 months after treatment, early biomarkers predicting ESKD or death are lacking. Recent studies encompassing not only LN have highlighted the central role of the alternative complement pathway (ACP), with or without histological evidence of thrombotic microangiopathy (TMA), as a key promotor of renal death., Methods: We assessed whether persistent isolated C3 hypocomplementemia (PI-LowC3), that is not accompanied by C4 hypocomplementemia, 6 months after kidney biopsy, is associated with an increased risk of death or ESKD in proliferative LN., Results: We retrospectively followed-up 197 patients with proliferative LN (51 with PI-LowC3) for a median of 4.5 years (interquartile-range: 1.9-9.0), 11 of whom died and 22 reached ESKD. After adjusting for age, gender, ethnicity, hypertension, mycophenolate, or cyclophosphamide use, PI-LowC3 was associated with a hazard ratio [HR] of the composite outcome ESKD or death of 2.46 (95% confidence interval [CI]: 1.22-4.99, P  = 0.012). These results were confirmed even after controlling for time-varying estimated glomerular filtration rate (eGFR) measurements in joint longitudinal-survival multiple regression models. After accounting for the competing risk of death, PI-LowC3 patients showed a strikingly increased risk of ESKD (adjusted HR 3.41, 95% CI: 1.31-8.88, P  = 0.012)., Conclusion: Our findings support the use of PI-LowC3 as a low-cost readily available biomarker, allowing clinicians to modify treatment strategies early in the course of disease and offering a rationale for complement blockade trials in this particularly at-risk subgroup of LN patients., (© 2022 International Society of Nephrology. Published by Elsevier Inc.)

Background: Image-based machine learning tools hold great promise for clinical applications in pathology research. However, the ideal end-users of these computational tools (e.g., pathologists and biological scientists) often lack the programming experience required for the setup and use of these tools which often rely on the use of command line interfaces., Methods: We have developed Histo-Cloud , a tool for segmentation of whole slide images (WSIs) that has an easy-to-use graphical user interface. This tool runs a state-of-the-art convolutional neural network (CNN) for segmentation of WSIs in the cloud and allows the extraction of features from segmented regions for further analysis., Results: By segmenting glomeruli, interstitial fibrosis and tubular atrophy, and vascular structures from renal and non-renal WSIs, we demonstrate the scalability, best practices for transfer learning, and effects of dataset variability. Finally, we demonstrate an application for animal model research, analyzing glomerular features in three murine models., Conclusions: Histo-Cloud is open source, accessible over the internet, and adaptable for segmentation of any histological structure regardless of stain., Competing Interests: Competing interestsJ.E.Z. is a paid consultant for Leica Biosystems. The remaining authors declare no competing interests., (© The Author(s) 2022.)

Background: COVID-19 can cause some individuals to experience chronic symptoms. Rates and predictors of chronic COVID-19 symptoms are not fully elucidated., Objective: To examine occurrence and patterns of post-acute sequelae of SARS-CoV2 infection (PASC) symptomatology and their relationship with demographics, acute COVID-19 symptoms and anti-SARS-CoV-2 IgG antibody responses., Methods: A multi-stage observational study was performed of adults (≥18 years) from 5 US states. Participants completed two rounds of electronic surveys (May-July 2020; April-May 2021) and underwent testing to anti-SARS-CoV-2 nucleocapsid protein IgG antibody testing. Latent Class Analysis was used to identify clusters of chronic COVID-19 symptoms., Results: Overall, 390 adults (median [25%ile, 75%ile] age: 42 [31, 54] years) with positive SARS-CoV-2 antibodies completed the follow-up survey; 92 (24.7%) had ≥1 chronic COVID-19 symptom, with 11-month median duration of persistent symptoms (range: 1-12 months). The most common chronic COVID-19 symptoms were fatigue (11.3%), change in smell (9.5%) or taste (5.6%), muscle or joint aches (5.4%) and weakness (4.6%). There were significantly higher proportions of ≥1 persistent COVID-19 symptom (31.5% vs. 18.6%; Chi-square, P = 0.004), and particularly fatigue (15.8% vs. 7.3%, P = 0.008) and headaches (5.4% vs. 1.0%, P = 0.011) in females compared to males. Chronic COVID-19 symptoms were also increased in individuals with ≥6 acute COVID-19 symptoms, Latent class analysis revealed 4 classes of symptoms. Latent class-1 (change of smell and taste) was associated with lower anti-SARS-CoV-2 antibody levels; class-2 and 3 (multiple chronic symptoms) were associated with higher anti-SARS-CoV-2 antibody levels and more severe acute COVID-19 infection., Limitations: Ambulatory cohort with less severe acute disease., Conclusion: Individuals with SARS-CoV-2 infection commonly experience chronic symptoms, most commonly fatigue, changes in smell or taste and muscle/joint aches. Female sex, severity of acute COVID-19 infection, and higher anti-SARS-CoV-2 IgG levels were associated with the highest risk of having chronic COVID-19 symptoms., Competing Interests: The authors have declared that no competing interests exist.

Pathogenic autoreactive antibodies that may be associated with life-threatening coronavirus disease 2019 (COVID-19) remain to be identified. Here, we show that self-assembled genome-scale libraries of full-length proteins covalently coupled to unique DNA barcodes for analysis by sequencing can be used for the unbiased identification of autoreactive antibodies in plasma samples. By screening 11,076 DNA-barcoded proteins expressed from a sequence-verified human ORFeome library, the method, which we named MIPSA (for Molecular Indexing of Proteins by Self-Assembly), allowed us to detect circulating neutralizing type-I and type-III interferon (IFN) autoantibodies in five plasma samples from 55 patients with life-threatening COVID-19. In addition to identifying neutralizing type-I IFN-α and IFN-ω autoantibodies and other previously known autoreactive antibodies in patient plasma, MIPSA enabled the detection of as yet unidentified neutralizing type-III anti-IFN-λ3 autoantibodies that were not seen in healthy plasma samples or in convalescent plasma from ten non-hospitalized individuals with COVID-19. The low cost and simple workflow of MIPSA will facilitate unbiased high-throughput analyses of protein-antibody, protein-protein and protein-small-molecule interactions., (© 2022. The Author(s), under exclusive licence to Springer Nature Limited.)

In our continuing examination of the role of exposomes in autoimmune disease, we use this review to focus on pathogens. Infections are major contributors to the pathophysiology of autoimmune diseases through various mechanisms, foremost being molecular mimicry, when the structural similarity between the pathogen and a human tissue antigen leads to autoimmune reactivity and even autoimmune disease. The three best examples of this are oral pathogens, SARS-CoV-2, and the herpesviruses. Oral pathogens reach the gut, disturb the microbiota, increase gut permeability, cause local inflammation, and generate autoantigens, leading to systemic inflammation, multiple autoimmune reactivities, and systemic autoimmunity. The COVID-19 pandemic put the spotlight on SARS-CoV-2, which has been called "the autoimmune virus." We explore in detail the evidence supporting this. We also describe how viruses, in particular herpesviruses, have a role in the induction of many different autoimmune diseases, detailing the various mechanisms involved. Lastly, we discuss the microbiome and the beneficial microbiota that populate it. We look at the role of the gut microbiome in autoimmune disorders, because of its role in regulating the immune system. Dysbiosis of the microbiota in the gut microbiome can lead to multiple autoimmune disorders. We conclude that understanding the precise roles and relationships shared by all these factors that comprise the exposome and identifying early events and root causes of these disorders can help us to develop more targeted therapeutic protocols for the management of this worldwide epidemic of autoimmunity.

Introduction: Podocyte depletion is a histomorphologic indicator of glomerular injury and predicts clinical outcomes. Podocyte estimation methods or podometrics are semiquantitative, technically involved, and laborious. Implementation of high-throughput podometrics in experimental and clinical workflows necessitates an automated podometrics pipeline. Recognizing that computational image analysis offers a robust approach to study cell and tissue structure, we developed and validated PodoCount (a computational tool for automated podocyte quantification in immunohistochemically labeled tissues) using a diverse data set., Methods: Whole-slide images (WSIs) of tissues immunostained with a podocyte nuclear marker and periodic acid-Schiff counterstain were acquired. The data set consisted of murine whole kidney sections ( n  = 135) from 6 disease models and human kidney biopsy specimens from patients with diabetic nephropathy (DN) ( n  = 45). Within segmented glomeruli, podocytes were extracted and image analysis was applied to compute measures of podocyte depletion and nuclear morphometry. Computational performance evaluation and statistical testing were performed to validate podometric and associated image features. PodoCount was disbursed as an open-source, cloud-based computational tool., Results: PodoCount produced highly accurate podocyte quantification when benchmarked against existing methods. Podocyte nuclear profiles were identified with 0.98 accuracy and segmented with 0.85 sensitivity and 0.99 specificity. Errors in podocyte count were bounded by 1 podocyte per glomerulus. Podocyte-specific image features were found to be significant predictors of disease state, proteinuria, and clinical outcome., Conclusion: PodoCount offers high-performance podocyte quantitation in diverse murine disease models and in human kidney biopsy specimens. Resultant features offer significant correlation with associated metadata and outcome. Our cloud-based tool will provide end users with a standardized approach for automated podometrics from gigapixel-sized WSIs., (© 2022 International Society of Nephrology. Published by Elsevier Inc.)

Sodium-glucose co-transporters (SGLTs) serve to reabsorb glucose in the kidney. Recently, these transporters, mainly SGLT2, have emerged as new therapeutic targets for patients with diabetes and kidney disease; by inhibiting glucose reabsorption, they promote glycosuria, weight loss, and improve glucose tolerance. They have also been linked to cardiac protection and mitigation of liver injury. However, to date, the mechanism(s) by which SGLT2 inhibition promotes systemic improvements is not fully appreciated. Using an obese TallyHo mouse model which recapitulates the human condition of diabetes and nonalcoholic fatty liver disease (NAFLD), we sought to determine how modulation of renal glucose handling impacts liver structure and function. Apart from an attenuation of hyperglycemia, Empagliflozin was found to decrease circulating triglycerides and lipid accumulation in the liver in male TallyHo mice. This correlated with lowered hepatic cholesterol esters. Using in vivo MRI analysis, we further determined that the reduction in hepatic steatosis in male TallyHo mice was associated with an increase in nuchal white fat indicative of "healthy adipose expansion". Notably, this whitening of the adipose came at the expense of brown adipose tissue. Collectively, these data indicate that the modulation of renal glucose handling has systemic effects and may be useful as a treatment option for NAFLD and steatohepatitis.

Genetic coding variants in APOL1, which encodes apolipoprotein L1 (APOL1), were identified in 2010 and are relatively common among individuals of sub-Saharan African ancestry. Approximately 13% of African Americans carry two APOL1 risk alleles. These variants, termed G1 and G2, are a frequent cause of kidney disease - termed APOL1 nephropathy - that typically manifests as focal segmental glomerulosclerosis and the clinical syndrome of hypertension and arterionephrosclerosis. Cell culture studies suggest that APOL1 variants cause cell dysfunction through several processes, including alterations in cation channel activity, inflammasome activation, increased endoplasmic reticulum stress, activation of protein kinase R, mitochondrial dysfunction and disruption of APOL1 ubiquitinylation. Risk of APOL1 nephropathy is mostly confined to individuals with two APOL1 risk variants. However, only a minority of individuals with two APOL1 risk alleles develop kidney disease, suggesting the need for a 'second hit'. The best recognized factor responsible for this 'second hit' is a chronic viral infection, particularly HIV-1, resulting in interferon-mediated activation of the APOL1 promoter, although most individuals with APOL1 nephropathy do not have an obvious cofactor. Current therapies for APOL1 nephropathies are not adequate to halt progression of chronic kidney disease, and new targeted molecular therapies are in clinical trials., (© 2022. Springer Nature Limited.)